Quantum Effects in Adenosylcobalamin-dependent Enzymes by M. Hossein Khalilian Boroujeni B.Sc., Chemistry, Razi University, 2014 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE in THE COLLEGE OF GRADUATE STUDIES (Chemistry) THE UNIVERSITY OF BRITISH COLUMBIA (Okanagan) April 2019 © M. Hossein Khalilian Boroujeni, 2019 The following individuals certify that they have read, and recommend to the College of Graduate Studies for acceptance, a thesis/dissertation entitled: Quantum Effects in Adenosylcobalamin-dependent Enzymes submitted by M. Hossein Khalilian Boroujeni in partial fulfillment of the requirements for the degree of Master of Science Examining Committee: Gino A. DiLabio, I. K. Barber School of Arts & Sciences Supervisor W. Stephen McNeil, I. K. Barber School of Arts & Sciences Supervisory Committee Member Kirsten Wolthers, I. K. Barber School of Arts & Sciences Supervisory Committee Member Michael Deyholos, I. K. Barber School of Arts & Sciences University Examiner ii Abstract The ability of radical enzymes to maintain tight control over the high reactive radical intermediates generated in their active sites is not completely understood. In this thesis, we report on a strategy that radical (B12-dependent) enzymes appear to exploit in order to manipulate and control the reactivity of one of their radical intermediate (5'-deoxyadenosyl radical) contained in the active site. The results of quantum mechanical calculations suggest that these enzymes utilize the little known quantum Coulombic effect (QCE), which causes the radical to acquire an electronic structure that contradicts the Aufbau Principle. This effect causes the energy of the singly-occupied molecular orbital (SOMO) of the radical to be well below that of the highest-occupied molecular orbital (HOMO), which renders the radical less reactive. The dynamic nature of the enzyme and its structure is expected to be such that the reactivity of the radical is not restored until it is moved into close proximity of the target substrate. It was found that the enzyme modulates the magnitude of the QCE and consequently reactivity through elaborate manipulation of the hydrogen bonding between 5'-deoxyadenosyl radical and nearby conserved glutamate residue. To the best of our knowledge, this work is the first study suggesting that both classical and quantum electrostatic factors contribute to both the catalytic power of these enzymes, and to the control of the reactivity and selectivity of the radical intermediates. In addition to B12 enzymes this electrostatic paradigm may be employed by other radical enzymes to attain selectivity of hard to control radical reactions. iii Lay Summary Radical enzymes are a class of proteins that use highly reactive intermediates known as radicals to perform challenging reactions. Exploiting these reactive intermediates requires a sophisticated control mechanism by the enzyme to prevent deleterious side reactions that can potentially damage the enzyme itself. The results of quantum mechanical calculations indicate that the enzymes we studied may use a sophisticated mechanism to control the reactivity of the radicals. This mechanism involves the interaction between charged groups in the enzyme active site and the radical through the quantum Coulombic effect (QCE), and tuning of the QCE through hydrogen bonding. The use of QCE by radical enzymes may be a general phenomenon. This thesis is the first report of the operation of QCE in enzyme systems. iv Table of Contents Abstract ......................................................................................................................................... iii Lay Summary ............................................................................................................................... iv Table of Contents ...........................................................................................................................v List of Tables ................................................................................................................................ ix List of Figures .................................................................................................................................x List of Symbols and Abbreviation ............................................................................................ xix Acknowledgements ................................................................................................................... xxii Dedication ................................................................................................................................. xxiii Chapter 1: Introduction ................................................................................................................1 1.1 Radical Enzymes ............................................................................................................. 1 1.2 Cobalamin-dependent Enzymes ...................................................................................... 2 1.2.1 General aspects ........................................................................................................... 2 1.2.2 Classes of B12 Enzymes .............................................................................................. 6 1.2.3 Isomerases ................................................................................................................... 8 1.2.4 Subgroups of Isomerases .......................................................................................... 10 1.2.4.1 Carbon-skeleton Mutases .................................................................................. 12 1.2.4.2 Aminomutases................................................................................................... 14 1.3 Catalytic Power in B12 Enzymes ................................................................................... 17 1.3.1 Hypotheses ................................................................................................................ 17 1.3.1.1 Strain Hypothesis .............................................................................................. 18 1.3.1.2 Electrostatic Effect ............................................................................................ 19 v 1.3.2 Selectivity in B12 Enzymes ....................................................................................... 22 1.4 Research Questions and Hypothesis ............................................................................. 24 Chapter 2: Orbital Conversion and Quantum Coulombic Effect ...........................................26 2.1 What is Orbital Conversion? ......................................................................................... 26 2.1.1 Orbital Conversion and Radical Stability ................................................................. 30 2.2 The Origin of the Orbital Conversion. Classical or Quantum Effect? .......................... 32 2.3 The Possible Role of QCE in B12 Enzymes .................................................................. 35 Chapter 3: Methods .....................................................................................................................37 3.1 Objective ....................................................................................................................... 37 3.2 Quantum Mechanic/Molecular Mechanic Simulations ................................................ 37 3.2.1 Background and Theory ............................................................................................ 37 3.3 QM/MM Calculations Details....................................................................................... 39 3.4 Molecular Orbital Energy Diagrams ............................................................................. 41 3.5 Transition State Calculations ........................................................................................ 42 3.6 BDE and RSE Calculations .......................................................................................... 43 3.7 Density State Plots (DOS) ............................................................................................ 43 3.8 Multiconfiguration Self-Consistent Field Theory ......................................................... 44 3.8.1 Background and Theory ............................................................................................ 44 Chapter 4: QM/MM Calculations Reveal the Presence of QCE in GM and MCM Enzymes ........................................................................................................................................49 4.1 Objective ....................................................................................................................... 49 4.2 Methodology (GLM Enzyme) ...................................................................................... 49 4.2.1 Models and Calculation Setup .................................................................................. 49 vi 4.2.2 QM/MM Calculations ............................................................................................... 54 4.3 QCE in GLM Enzymes ................................................................................................. 55 4.3.1 Orbital Configurations and Energies Obtained from QM/MM Calculations ........... 55 4.3.2 Analysis and Discussion ........................................................................................... 61 4.4 Methodology